Biting flies and Trypanosoma vivax infection in three highland districts bordering lake Tana, Ethiopia

Veterinary Parasitology 142 (2006) 35–46 www.elsevier.com/locate/vetpar

Biting flies and Trypanosoma vivax infection in three highland districts bordering lake Tana, Ethiopia A. Sinshaw a, G. Abebe b,*, M. Desquesnes c,d, W. Yoni c a b

Bureau of Agriculture, Amhara National Regional State, P.O. Box 437, Bahir Dar, Ethiopia Faculty of Veterinary Medicine, Addis Ababa University, P.O. Box 34, Debre Zeit, Ethiopia c CIRDES, BP 454 Bobo-Dioulasso, Burkina Faso d CIRAD-EMVT, Campus International de Baillarguet, 34398 Montpellier, France Received 6 March 2006; received in revised form 13 June 2006; accepted 21 June 2006

Abstract An epidemiological study was conducted to determine the prevalence of trypanosomosis in cattle, small ruminants and Equidae, and to identify biting flies; potential mechanical vectors of trypanosomes in the three districts of Bahir Dar Zuria, Dembia and Fogera, bordering lake Tana, Ethiopia. About 1509 cattle, 798 small ruminants and 749 Equidae were bled for the prevalence study using the buffy-coat method and the measurement of the hematocrit value. Sixty-six NGU and 20 monoconical traps were deployed for the fly survey. The results indicated the presence of trypanosomes in 6.1% (92/1509) of the cattle with a maximum during the late rainy season (9.6%) than the early dry season (3.6%) at Fogera district. Prevalence at the district level varied from 4% to 9.6%. Only one sheep (1/122) and one goat (1/676) were found positive for T. vivax-like trypanosomes and none of the Equidae was positive. All the trypanosomes encountered in cattle belong to the single species of T. vivax. The PCV was negatively associated with detection of T. vivax (21.6% in infected versus 25.4% in non-infected cattle). A total of 55,398 biting flies were caught of which 49,353 (89.08%) belong to Stomoxys, 4715 (8.51%) to horse flies and 1330 (2.4%) to Chrysops species. There was no tsetse fly. Species identification has indicated the presence of Atylotus agrestis, Chrysops streptobalia, Stomoxys calcitrans, S. nigra, S. pulla, S. pallida, S. sitiens, S. taeniata, S. uruma, Haematopota lasiops and Hippobosca variegata. The overall apparent density was 214.7 flies/trap/day. Seasonal comparison showed higher fly catches in the late rainy season than the early dry season. This study indicated that T. vivax infections culminate in cattle at the same time as mechanical vectors such as Stomoxys sp. and Atylotus agrestis. Therefore, attention towards T. vivax infection in cattle is essential to control the impact of the disease on productivity. A further study on biting flies is recommended. # 2006 Elsevier B.V. All rights reserved. Keywords: Biting flies; Equidae; Ethiopia; Lake Tana; Trypanosoma vivax; Ruminants

1. Introduction Though, the role of mechanical vectors in the transmission of African livestock trypanosomes has always been controversial relative to tsetse flies, their

* Corresponding author. E-mail address: [email protected] (G. Abebe). 0304-4017/$ – see front matter # 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.vetpar.2006.06.032

cyclical vectors, in recent experimental works it was successfully demonstrated that mechanical transmission of T. vivax to cattle was effected by African tabanids with high incidence rates of 63% by Atylotus agrestis (Desquesnes and Dia, 2003) and 75% with Atylotus fuscipes (Desquesnes and Dia, 2004) within 20 days. Ethiopia, as part of the African continent shares a substantial loss from trypanosomosis. Apart from the cyclical transmission of trypanosomosis by the Glossina

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species, it is highly considered that mechanical transmission is a potential threat to livestock productivity in some parts of Ethiopia (Abebe and Jobre, 1996). However, information on prevalence of non-cyclically transmitted trypanosomosis in domestic animals, and the vectors involved in Ethiopia is scanty and sufficient data in a compiled form is not available. Therefore, this study was undertaken with the objectives of investigating the distribution of mechanically transmitted trypanosomosis of domestic animals (cattle, small ruminants, and equines) and assessing the distribution of mechanical vectors (biting flies) and identify at a species level in three districts adjacent to lake Tana, Amhara National Regional State (ANRS), Ethiopia. 2. Materials and methods 2.1. Study area and animals The study was undertaken in three discrete districts bordering lake Tana, namely Bahir Dar Zuria, Fogera, and Dembia; located in West Gojjam and South Gonder and North Gonder administrative Zones of the Amhara National Regional State, respectively, Ethiopia (Fig. 1). The study districts are densely populated and most of the land is intensively cultivated and in the rainy season, particularly in the so-called Fogera plain, most of the cultivable land will be water logged. Cattle, small ruminants and Equidae were used as study animals. Among the domestic animals, cattle are the dominant species raised and the Fogera cattle breed/type is concentrated in the study districts.

Fig. 1. Map of Ethiopia showing study districts around lake Tana. The source of the original map is not available and the word Dembya is used as the name has both slangs in local language.

2.2. Study design, sample collection and identification The study was based on cross-sectional type of investigation. A combination of stratified, multistage and purposive sampling methods were applied according to Toma et al. (1999) and Putt et al. (1988). The three discrete study districts were selected from three zones of the ANRS (first stage) to represent areas bordering lake Tana. Then, a list of Peasant Associations (PAs) within districts were compiled from a data obtained in the district’s agricultural office (second stage) and sampling PAs were selected based on representation of the respective districts and accessibility. Villages were selected in collaboration with the respective district’s animal health personnel, selected by purposive sampling on the basis of prior information on the problem, farmer’s co-operation, logistics, share of communal grazing land and accessibility (third stage). Selected villages and herds grazing within the same grazing land were considered as strata. With in each stratum, sampling was performed irrespective of the other strata. Then representative numbers of animals (considering sex and age) were sampled. Representative numbers of animals were sampled from each village. Parameters like age, sex, body conditions score and reproductive status (parity, lactation, pregnancy, and abortion) were recorded for each individual animal. Individual animals with age greater than 6 months were considered. To determine the sample size, a trypanosomosis prevalence rate of 10% (the average prevalence for cattle in West Gojjam and South and North Gonder regions) was taken into consideration. Hence, for cattle, 900 samples (10% expected prevalence, 95% confidence level and 2% precision) were taken. However, in case of small ruminants and equines since there was no information available about prevalence estimates for these species, 20% prevalence rate was used to estimate the sample size. Therefore, a sample size of 711 small ruminants, (20% expected prevalence, 95% confidence level and 3% precision) and 711 Equidae among which 608 donkeys and 141 mules (20% expected prevalence, 95% confidence level and 3% precision) were assumed, respectively (Putt et al., 1988; Toma et al., 1999). Therefore, a total number of 2322 domestic animals were required for the study. However, a total of 1509 cattle (at Fogera 592 samples in the late rainy season and 609 in the early wet season, 125 at Dembia and 133 at Bahir Dar Zuria), 798 small ruminants and 749 equines were practically considered. In case of cattle, the comparison of two seasons was considered (late rainy and early dry) at the Fogera district that makes the

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total sample size 1509. Hence, a total of 3056 animals were sampled for this study. 2.3. Parasitological and hematological examinations Animals were bled from the peripheral ear vain using microhematocrit capillary tubes for the purpose of parasitological examination of trypanosomes, estimation of anemia and species identification. A microhematocrit capillary tube containing 70 ml of blood were centrifuged for 5 min using the Hawksley microhematocrit centrifuge, the PCV was read for estimation of anemia using a hematocrit reader and the buffy-coat examination done as described in Murray et al. (1983). For the purpose of species identification, a thin blood smear was prepared from the buffy-coat for those samples that were positive on buffy-coat examination and stained with Giemsa stain and examined under a microscope using the oil immersion 100 objective (Murray et al., 1983). 2.4. Survey of flies From October 2003 to February 2004, a total of 86 (63 during late rainy season and 23 during early dry season) standard traps developed for tsetse fly trapping (66 NGU, and 20 monoconical) (Drees and Jackman, 1998) were deployed in the three districts. All the traps were baited uniformly with octenol (1-oct-3-nel) and acetone. The poles of traps were greased to prevent fly predators mainly ants. Traps were allowed to stay at the site of deployment for a period of 72 h before collection. Trap deployment sites were selected to represent all habitats that could be related to fly multiplication, behavior, feeding, and other related aspects. Hence grazing lands, cattle barns, swampy areas, bushy areas, riverbanks, watering points and animal congregation sites were purposely included and extent of such habitats were recorded represented by numbers to be transformed into percentage values expressing the level of the selected site of deployment later for analysis. After 72 h of deployment, the catchments of each trap was sorted by fly’s tribe, and then counted. The cover of a new matchbox were labeled by district, PA, trap type, date, site description, name of fly tribe, and other informations that may be relevant from the aspect of data analysis was included. Then, fly samples were put in the matchbox for a further species identification (Murray et al., 1983). Flies collected from traps and stored in a clean matchbox were identified at a species level at the Faculty of Veterinary Medicine, Addis Ababa University and representative samples were sent to South Africa and Burkina Faso (CIRDES) for simulta-

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neous identification by entomologists and experts in veterinary parasitology/entomology as recommended by Drees and Jackman (1998). Flies were mounted on a stereomicroscope in the Department of Parasitology Faculty of Veterinary Medicine; Addis Ababa University and species identification was done according to Oldroyd (1952a,b, 1954, 1957). After identification of the samples at a species level, flies were permanently preserved in a 70% ethyl alcohol (Drees and Jackman, 1998). 2.5. Data analysis Data on individual animals, parasitological examination results, and data on entomology was inserted in to MS Excel Spread Sheets Program (Microsoft Corp.) to create a database and transferred to the STATA and SPSS software programs of the computer before analysis. Descriptive statistics, confidence interval, Student’s t-test, Pearson’s correlation, and ANOVA were used to express results and compare variables. The Intercooled STATA 7 (STATA, 2001) and SPSS (SPSS, 2002) software of the Computer Program were applied for the statistical analysis. The prevalence rate of trypanosome infection was calculated as the number of parasitologically positive animals as examined by the buffy-coat method (Murray et al., 1983) divided by the total number of animals investigated at that particular time. Confidence intervals (95%) for the PCV of trypanosome-infected and noninfected and among different physiological parameters (lactation, sex and pregnancy) were calculated. ANOVA was used to compare the prevalence rates of trypanosome infections in different districts, peasant associations, and seasons (Intercooled STATA, 2001 and SPSS, 2002). Student’s t-test was utilized to compare the mean PCVof the parasitic animals with that of the aparasitemic animals (Intercooled STATA, 2001 and SPSS, 2002). For the data on fly survey since the number of flies caught varied widely, the data was transformed to a logarithmic scale using the transformation y = ln (x + 1) before the statistical analysis. Then, Student’s t-test was used to compare the difference of fly catch between the NGU and monoconical traps and between the two seasons. ANOVA was applied to compare the mean fly catch difference among the different trap deployment sites. 3. Results 3.1. Results of the cross-sectional study Out of a total of 1509 cattle sampled the overall rate of positive was 6.1% (92/1509). Prevalence was significantly ( p < 0.001.) higher during the late rainy season

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Table 1 Parasitological prevalence of Trypanosoma vivax in cattle of the three districts Districts

Season

Number of cattle

Prevalence (%)

Examined Positive Fogera

Early wet season Late dry season

592 609

57 22

9.6 3.6

Dembia

Early wet season

175

7

4.0

Bahir dar Early wet season zuria

133

6

4.5

1509

92

6.1

Total

Both seasons

9.6% (57/592) than the early dry season 3.6% (22/609) at Fogera district where the two seasons were compared. Prevalence at district level in early wet season has significantly ( p < 0.005) varied from 9.6% (57/592) at Fogera district to 4.5%(6/133) at Bahir Dar and 4% (7/175) at Dembia (Table 1). In cattle all the trypanosomes encountered belong to the single species of T. vivax. However, the Trypanosoma species in sheep and goats, though seemed T. vivax from the buffy-coat movements, could not be confirmed by Giemsa-stained buffy-coat smear observation due to the very low number of parasites. In cattle, the variation in prevalence of T. vivax with regard to age and sex was not statistically significant ( p > 0.05). Of 798 small ruminants (122 sheep and 676 goats), only 1 sheep 1/122 (0.82%) and 1 goat, 1/676 (0.15%), were found positive for Trypanosoma. None of the 608 donkeys and 141 mules was positive for trypanosomes. 3.2. Results of the PCV values and productive parameters

Fig. 3. Body condition score in relation to T. vivax infection and PCV in cattle.

vivax-positive and -negative animals, respectively. The difference between the two groups is 3.8%. ANOVA performed for the districts has indicated that the PCV of cattle was with in the normal range for the species (Fig. 2) but there was a significant difference ( p < 0.001) in mean PCV of the three districts (Bahir Dar Zuria, 25.2% CI = 24.6–25.8%, Dembia 27.4% CI = 26.9–27.8% and Fogera 25.8% CI = 25.5–26.1%). Association of PCV (n = 1509) with body condition score, and presence of T. vivax infection using Pearson’s correlation indicated that PCV was positively related with body condition score (BCS) (r = 0.123, p < 0.05), and negatively related with presence of T. vivax infection (r = 0.221, p < 0.05). A negative relationship was also observed between BCS and T. vivax infection (r = 0.138, p < 0.05) (Fig. 3). There was also a highly significant ( p < 0.0001) variation in PCV between the late rainy season and early dry season (Fig. 4).

The PCV of cattle was significantly ( p < 0.001) affected by T. vivax infection and it was 21.6% (95% CI = 20.9–22.3) and 25.4% (95% CI = 20.9–22.3) in T.

Fig. 2. PCV profiles in cattle of the three study districts.

Fig. 4. PCV of cattle in the two seasons at Fogera district. S.E.M. = 25.2  0.1 (late rainy season), 24.3  0.2.

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Table 2 PCV of female cattle (4 years) categorized by reproductive status Reproductive parameters

Reproductive status

No. of animals

Mean PCV

95% CI

Lactation

Non-lactating Lactating

347 415

26.5 25.8

26.0–27.0 25.5–26.2

Pregnancy

Non-pregnant Pregnant

661 101

25.9 27.9

25.6–26.2 27.1–28.7

Abortion

No abortion Aborted

729 33

26.2 25.7

25.9–26.5 24.4–27.0

Table 3 Relationship among the reproductive and productive indices in female cattle 4 years of age BCS BCS Parity Pregnancy Lactation PCV Infection (T. vivax)

Parity

Pregnancy

Lactation 0.162

0.162** 0.238** 0.072*

0.123**

0.163**

**

PCV

Infection (T. vivax) **

0.238 0.123** 0.163** 0.079*

0.079*

0.072*

0.186** 0.186**

**Correlation is significant at the 0.01 level. *Correlation is significant at the 0.05 level. Only significant values are presented (n = 762, Pearson’s correlation).

3.3. Results of the fly survey

lected by hand from the body of cattle at Dembia district and this fly genus was not observed to enter in any one of the two trap types. From the representative samples subjected for species identification, Atylotus agrestis (4111), Chrysops streptobalia (688), Stomoxys nigra (293), S. calcitrans (64), S. sitiens (45), S. pulla (38), S. taeniata (18), S. uruma (21), and S. pallida (1), Haematopota lasiops (1) and Hippobosca variegata (8) were confirmed at a species level to be present in the three study districts. There was a variation in percentage distribution of Stomoxys species identified (Fig. 6). No tsetse fly was captured in any of the study districts. Among the horse flies, A. agrestis was the most abundant and all the Chrysops belong the species Chrysops streptobalia. Seasonal comparison of fly

From 86 traps deployed at the three study districts, a total of 55,398 flies were caught. Of which 49,353 (89.08%) belong to the family Stomoxys, 4715 horse flies (8.51%) and 1330 (2.4%) Chrysops. The number of fly tribes collected was significantly different ( p < 0.001) for each fly category, being in the order of Stomoxys (89.1%), horse flies (8.5%), and Chrysops (2.4%) (Fig. 5). The overall apparent density of flies was 214.7 flies/trap/day. Results on fly survey in this study has revealed the presence of various fly tribes and genera including horse flies, Stomoxys, Chrysops, Hematopota and Hippobosca. Hippobosca was col-

Fig. 5. The relative percentage distribution of total fly catch. S.E.M. of total fly catch = Stomoxys (49,353  0.5), horse flies (4715  0.5), Chrysops (1330  0.5).

Analysis of variance and Pearson’s correlation statistics for productive and reproductive indices in female cattle (n = 762) of age > 4 (females at reproductive age) were performed to predict and relate PCV with other parameters, respectively. By taking PCV as an outcome variable using ANOVA has revealed that PCV was significantly negatively affected by parity ( p < 0.01), and lactation ( p < 0.05). However, body condition score ( p < 0.001) and pregnancy ( p < 0.01) had a positive impact on PCV. Though abortion had lowered the PCV values, there was no statistical difference ( p > 0.05) with those without abortion (Tables 2 and 3).

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Fig. 6. The percentage distribution of Stomoxys species identified.

Fig. 7. Distribution of flies catches during the late rainy and early dry seasons. It indicates the total number of seasonal fly catches per type of fly genera.

Fig. 8. Comparison of flies trapped by the monoconical and NGU traps. It indicates the total number of fly catches for each trap type.

catches using 23 traps per season at Fogera district (17 NGU and 6 monoconical traps per season) indicated that there is a remarkably significant variation ( p < 0.0001) in overall fly genera number between the two seasons for horse flies, Chrysops and Stomoxys (Fig. 7). This seasonal variation was consistent when each trap type (either NGU or monoconical) was compared for each season. Comparison of the NGU and monoconical trap has indicated that the NGU trap had a significantly ( p < 0.05) high catch of horse flies than the monoconical where as the monoconical trap had a significantly high catch of Stomoxys ( p < 0.0001) over the NGU trap. But there was no significant difference in catchments between the two traps for Chrysops species (Fig. 8). Traps deployment sites were categorized and compared for farmers village, grazing land, swampy locations, cattle barns, dry sites, cattle congregation

Fig. 9. Fly catchments in relation to the level of swamp. 0: 0%; 2: 50%; 3: 75%; 4: 100%. Unable to retrieve original data.

A. Sinshaw et al. / Veterinary Parasitology 142 (2006) 35–46

Fig. 10. Fly catchments in relation to the level of dryness. 0: 0%; 2: 50%; 3: 75%; 4: 100%. Unable to retrieve original data.

areas, river banks, watering points, high way, and areas covered with vegetation. Comparison of each fly type by trap deployment sites using ANOVA has indicated that the catch of horse flies, catch of Chrysops, and Stomoxys ( p < 0.001) was significantly high in and around swampy areas and low at dry sites ( p < 0.001). All the other site variables included in the ANOVA model did not show any significant catch difference (Figs. 9 and 10). 4. Discussion The percentage prevalence of positive samples by microscopical examination of the buffy-coat for trypanosomes is within the range of other previous reports of studies conducted in neighboring and similar districts of this study and has varied from 2% to 16% (Getinet, 1994; Mihiret, 1995; Enyew and Abebe, 1997; Cherinet, 1999; Terefe and Abebe, 1999). The presence of variation in prevalence of T. vivax among the districts where there are no tsetse fly reports is already documented (Mihiret, 1995; Getinet, 1994; Terefe and Abebe, 1999; Aklilu, 2002). A prevalence study conducted in a similar and adjacent survey sites of this study by Hassen (1988), reported zero prevalence at Gayint, Wogera, Estie, Armachiho, Simada, and Debark to a 5% (3/60) at Kemkem, 5.75% (5/87) at Dera, 7.5% (6/80) at Dembia and 8.43% (22/261) at Fogera. Similarly, in a wide area survey conducted in Zambia, Sinyangwe et al. (2001) reported that prevalence in individual villages varied between 0% and 64%. A prevalence variation that laid between 0% and 43% has also been reported by Mwambo et al. (2001) in Tanzania. Such very high variations between farms are classically described in areas where T. vivax is strictly mechanically transmitted in cattle, such

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as in Colombia (Wells et al., 1982; Otte et al., 1988) and French Guyana (Desquesnes, 2004). This variation among districts in the present study could be attributed to the biting fly population and type present in each locality, which is dependent on microclimate, animal herd density, distance between herds and other various factors (Foil, 1996a); it is also a characteristic of mechanically transmitted cattle trypansomosis (Desquesnes, 2004). A significantly high rate of infection following the months with high rainfall reported in our study is due to the emergence of biting flies at a high rate as it has matched with the high number of biting flies collected. However, the decrease in prevalence in the early dry season would be complimented by the fact that owners take their animals for trypanocidal drug treatment and as the infected animal number decreases, the source of infection decreases. Hence, in concomitant with the decrease in fly population during the dry season, the prevalence of T. vivax is expected to decrease. The selfcure phenomenon following T. vivax infection would also contribute to the decrease in prevalence. This self-cure phenomenon is ascribed to the limited number of variable surface glycoprotein in T. vivax (Gardiner, 1989). Such seasonal difference in the prevalence of T. vivax infection rates was reported by adjacent areas of the present study districts (Getinet, 1994; Mihiret, 1995; Enyew and Abebe, 1997; Cherinet, 1999; Terefe and Abebe, 1999). In Nigeria, a study conducted on zebu cattle has revealed a higher infection rates during rains (9.3%) than in the dry season months (1.5%). In this area, tsetse were encountered at low density and T. vivax was the predominant species accounting 81% infections in the rainy season and 100% in the dry season (Kalu, 1996). This suggests that biting flies would mediate T. vivax infections when the tsetse fly density is either low or absent. Similarly, Tamasaukas et al. (1996) indicated that bovine trypanosomosis caused by T. vivax was low during part of the dry season and suggested that it is due to major changes in the environmental and agro-ecological conditions of the farms during this season in comparison with those observed in the rainy season. D’Ieteren et al. (1988), found that prevalence for cattle and sheep was two to four times lower in the dry season than the months with highest rain fall and Kalu and Lawani (1996), reported that infection rates doubled during the rainy season (7.6%) as compared with an average of 3.8% during the dry season. The very low prevalence in sheep and goats might be due to that biting flies prefer cattle than other domestic animals (Kniepert, 1981) and in a mixed farming system in the study areas where different species of animals are kept together in a communal grazing area, biting flies

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would preferably attack cattle, leaving most of the small ruminants uninfected. However, the presence of single infections in one sheep and one goat indicates that these species would get the infection and other diagnostic methods apart from the parasitological techniques might reveal the extent of the infection rate in these small stocks. As for horses, they may either remain uninfected due to a separated breeding, or due to a low level of sensitivity to the local stocks of T. vivax. Kniepert (1981) showed that the rate of attack by species of Tabanidae on cattle depended on the position, size and coloration of the host. Tabanid species studied preferred a particular site for feeding caused by variations in the length of the hair, thickness and tensile strength of the skin of the animal. The tabanids preferred sites in which these characters correlated with the absolute and relative length of their proboscis. A high prevalence rate in cattle and low in small ruminants has been reported by different workers previously. Defly et al. (1988) indicated that livestock species had a major effect on trypanosome prevalence. They found that prevalence in trypanotolerant cattle was three times than in sheep kept in the same area, but there was no difference between the trypanosome prevalence of sheep and goats. This difference in cattle and sheep was greatest where T. vivax causes about 90% of parasitemia in cattle and about 95% in sheep. Kalu and Lawani (1996), observed a prevalence of 5.3%, 1.2% and 0.7% in cattle, sheep, and goats, respectively. In Southwest Ethiopia, in a tsetse-infested area, Dinka and Abebe (2005) has reported a 5.1% (7.6% in sheep and 3.6% in goats) prevalence of trypanosomosis in small ruminants and the prevalence varied from village to village. Similarly, Hendy (1988), found a found a prevalence of 16% in east African short horn zebu. However, no parasites were found in goats (0/280) and only 4/172 sheep were found parasitemic. He proposed that there might be a problem in the diagnosis of disease. This may be a particular problem in goats and sheep, especially if some degree of trypanotolerance exists. Detection of trypanosomes in the blood may then be difficult. However, more sensitive/specific tests such as the PCR technique (Solano et al., 2001) would be applied to reaffirm our findings. Anosa et al. (1995), reported a parasitological prevalence of 4.5%, 2.7% and 2.2% in cattle, sheep and goats. Consideration of the presence of infection in small ruminants in the present study is important since they would act as a reservoir of infection for cattle. Desquesnes (1997) has skeptically put equines susceptibility to the Latin American T. vivax strain; however, in Senegal and Burkina Faso equines are regularly found infected by T. vivax. Perhaps,

though equines are preferred for attack by biting flies next to cattle, they might not be susceptible to the mechanically transmitted strain of T. vivax present around lake Tana. Results on hematological values reported in the present study where T. vivax was the only species of Trypanosoma encountered in cattle of the three districts, the degree of anemia as measured by the PCV was profound. Such significant difference in PCV of cattle due to trypanosomosis in ruminants is available in various works done so far and that of T. vivax infections is tabulated in the literature review part of this paper (Defly et al., 1988; D’Ieteren et al., 1988; Maloo et al., 1988; Mulatu et al., 1988; Ordner et al., 1988; Getinet, 1994; Mihiret, 1995; Abebe and Jobre, 1996; Kalu, 1996; Enyew and Abebe, 1997; Terefe and Abebe, 1999; Aklilu, 2002). Desquesnes and Dia (2003, 2004), demonstrated that hematocrit values of infected cattle (after experimental mechanical transmission of T. vivax with Atylotus agrestis and Atylotus fuscipes, respectively) has decreased during the infection period indicating the notable pathogenic effect of mechanically transmitted T. vivax and this transmission of T. vivax proved to be very efficient with an incidence observed of 75%, from an initial prevalence of 20% infected animals (donor animals). Taylor (1998) indicated that anemia persists during the chronic stages of infection when parasitemia is generally quite low, probably because different mechanisms are involved in its genesis during the acute and chronic stages of infection. This suggests that control of parasitemia and control of anemia is unrelated in the chronic phase when immune infections are depressed and anemia is sustained through dyserythropoiesis. In the present study, in female cattle (infected or non-infected) and goats at reproductive age, PCV was negatively associated with lactation and parity. Formerly, Ordner et al. (1988), found that one or more trypanosome parasitemic months detected in the cow during the breeding year depressed the average PCV in the same period by 4% units. They stated that, lactation status of the cow has significantly affected the average PCV level during the breeding year, which was 1.5% lower in lactating as compared to dry cows. Similarly, in sheep, the mean PCV of lactating ewes was lower than that of gestating ewes (D’Ieteren et al., 1988). PCV was positively associated with the body condition score of cattle and there was a low body condition score in T. vivax-infected animals than non-infected groups. This variation in PCV in relation to reproductive and productive indices would be important if considered with the aspect of predisposing factors to T. vivax infection and taking PCV as one major criteria of

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assessing Trypanosoma infection. T. vivax was found in different peasant associations in the three districts, and it seems that species (hosts) are important factors for the development of infection and the disease as mostly detected in cattle, negligible in small ruminants and nil in equines. In general, the present findings indicated that trypanosomosis due to T. vivax has established in the three districts in affecting cattle productivity and small ruminants are at a higher risk of infection and development of the disease in the three districts bordering lake Tana, Ethiopia. Findings of the fly survey in this study could be considered as a preliminary work. In Ethiopia, though there has not been a specific study on biting flies in alienation from the usual tsetse fly studies, few authors have reported the name of some of the biting flies as Tabanidae and Stomoxyinae at a family level (Enyew and Abebe, 1997; Kidane-Mariam, 2000). However, Kigaye and Jiffar (1991) in a survey of ectoparasites of cattle in Harar and Dire Dawa districts, south-Eastern part of Ethiopia have reported the presence of 10 species of ‘‘stable flies’’ Stomoxys calcitrans, S. nigra (S. niger), S. sitiens, S. varipes, S. bilineata (S. niger bilineatus), S. brunnipes, Lyperosia spinigera (Haematobia spinigera), Lyperosia minuta (H. minuta), L. thirouxi (H. hirouxi) and H. hirtifrons. Seven ‘‘housefly’’ species, one H. variegata, and two tabanids (Chrysops obliquefasciata and Hematopota atellicorne). However, various workers in different African countries have reported the presence of different biting and non-biting fly tribes/genera found in the present study. The most abundant Atylotus agrestis and Stomoxys species in our collection were previously reported from Sudan, Saudi Arabia, Nigeria, Burkina Faso, Mauritania and elsewhere (Adeyefa and Dipeolu, 1986; Leclercq, 1986; Amoudi and LeClercq, 1993; Amoudi and Leclercq, 1996; Dia et al., 1997, 1998). Acapovi et al. (2001) reported that out of 2471 Stomoxyinae captured in North of Coˆte d’Ivoire, 70.7% were Stomoxys niger and 29.3% S. calcitrans. The Stomoxyinae represented by two species only, made up about 45% of the biting flies captured. They will have to be considered when evaluating the impact of biting insects on cattle. This finding is similar to our reports that Stomoxyinae are present at a high population. In a study conducted in Mauritania, Dia et al. (1998) have reported a similar distribution of flies genera as has been found in this study (Tabanus, Atylotus, Stomoxyinae, and Hippobosca) and they have collected Hippobosca from the body of the animals as has been done in our study. In Burkina Faso, the distribution of fly species under natural condition was Atylotus agrestis 20%, A. fuscipes 4%, Chrysops distinctipennis 12%, Tabanus taeniola 2%, T.

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sufis 16%, and Stomoxys niger 46% (Desquesnes and Dia, 2003). In our study, the large count of biting flies during the rainy season collected using traps developed for tsetse fly trapping was complemented by the use of 1-octen-3-ol and acetone. Different authors confirmed that the use of attractants namely, 1-octen-3-ol, acetone, CO2, ammonia, phenols and cow urine-baited traps alone or in combination had improved the catch of Tabanidae than the non-baited traps (Hribar et al., 1992; Hayes et al., 1993; Foil and Hribar, 1995; McElligott and Lewis, 1996; Djiteye et al., 1998; McElligott and Lewis, 1998; Kristensen and Sommer, 2000; Ngare and Mwendia, 2001). In one experiment, Vale (1982) demonstrated that the catch of Stomoxyinae and non-biting Muscidae in the presence and absence of odor was 83.1% and 38.3%, respectively. The efficiency of the NGU trap over the monoconical in trapping the horse flies and the monoconical over the NGU for Stomoxys observed in this study was due to that trap design, and color are factors of attraction and larger flies prefer larger traps and vice versa. This efficiency of the NGU trap over the monoconical in trapping the horse flies has been previously observed by different workers (Amsler et al., 1994; Foil and Hribar, 1995; Djiteye et al., 1998). The NG-2G and F3 traps and the screen-trap were significantly more effective than the biconical and monoconical ones in having a high catch of Tabanidae. The large screen produced relatively small catches of Stomoxyinae and non-biting Muscidae, where as the small screen produced relatively large catches of these flies (Vale, 1982). The population peak of most species, during the late rainy season including those with higher vector potential, suggests that the rainy season can be considered as the period of potentially higher risk of mechanical transmission of pathogens by biting flies. This high population density of biting flies recorded in this study at the end of the rainy season is due to that biting flies requires a wet habitat for multiplication and larval growth is also dependent up on wet soil/mud. High population density of various biting flies following the rains is reported by various workers in different countries (McElligott and Galloway, 1991; Gorayeb, 1993; Cilek et al., 1994; Dia et al., 1997). Dia et al. (1997) found that species of Atylotus agrestis, Tabanus taeniola, T. sufis, Haematobia minuta and the hippoboscids (Hippobosca camelina and H. variegata) were particularly abundant during the end of the rainy season, but could be found throughout the year at a very low density. Similarly, Dia et al. (1998) in Mauritania, found that most of the Tabanidae were caught between October and November at the end of the rainy season.

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Gorayeb (1993) indicated that the correlation of certain climatic factors with the seasonal abundances of common tabanid species was investigated and significant relations were found for some species with air temperature, relative humidity of the air, rainfall, insulation, and evaporation potentiality or light intensity. Different ecological habitats where flies are numerous or scarce investigated so far indicated that each fly genera has adapted to a certain locality for breeding, feeding, resting and host seeking. Dia et al. (1998) reported that high number of Tabanidae was collected from an area with ample water and traps placed in the pasture near this watery area caught 80% of the population. Janzen and Hunter (1998) collected most of the Chrysops from a bog habitat. Lewis (1987) reported that the most consistently favorable collecting sites for Chrysops were natural or artificial ponds, which is agreement with what has been observed in our study. Lago and Testa (1990) reported that in an area surrounding tidal marshes Chrysops and Tabanus were the common horseflies. The largest numbers of tabanids were caught in the gallery and the fewest in the forest (Acapovi et al., 2001). Dia et al. (1997), found that in most of the time, T. taeniola and A. agrestis were caught in pastures, while T. sufis was caught by traps placed near water. Collection of adult flies is also dependent on larval habitat. Foil (1996a) reported that the larva of tabanids feed on organic debris and small invertebrates in a variety of aquatic to semi-aquatic habitats. However, stable fly larvae develop in manure-spilled feed and decaying vegetation. Cattle manure on cattle feed lots is an important medium for stable fly larval development. In general, the fly survey in this study demonstrated that there is a high fly density during the late rainy season and different biting fly genera/species. Therefore, apart from transmission of Trypanosoma, these biting pests are important in transmitting many other livestock diseases so far studied (Krinsky, 1976; Foil, 1989) and their economic importance in livestock productivity through loss of weight and condition (loss of blood, annoyance, predisposing to infection) should be considered together. Hollander and Wright (1980) estimated the blood loss in cattle caused by tabanids to be more than 200 ml/animal/day and Stomoxyinae and tabanids cause weight loss due to blood loss and annoyance as well as create feeding lesion sites which may promote contaminative transmission of agents or myiasis, (Foil, 1996a). The most abundant fly species Atylotus agrestis found in our study has been recently demonstrated as an effective mechanical vector of T. vivax at high rate of 63% (Desquesnes and Dia, 2003) and the authors concluded that in Africa, the

epidemiology of trypanosomosis in cattle is also tabanid dependent and the eradication of tsetse flies will not necessarily lead to eradication of T. vivax. 5. Conclusion In general, the present study indicated that trypanosomosis due to T. vivax is an important disease limiting cattle productivity and small ruminants are infected that may act as a source of infection for cattle. Infection with T. vivax has negatively affected the PCV and BCS of cattle. This indicates that T. vivax infection of cattle in the study area causes loss of body weight and production. The presence of various biting flies and the absence of tsetse flies in this investigation indicate that T. vivax infection in the study area is caused by mechanical transmission mediated by biting flies. The presence of biting flies at a higher density during the late rainy season and the concomitant higher prevalence of T. vivax in this same season supports that biting flies are the main epidemiological factors for T. vivax infection. The presence of biting flies in the early dry season at a small number would help to assure the continuity of T. vivax circulations in cattle herds at a low rate and when the next rainy season favors vector multiplication those circulating infections would flare up in a major proportion of the herd. Therefore, a particular attention towards T. vivax infection in cattle is essential to control the impact of the disease on cattle productivity. Development of control options that could minimize biting flies especially in seasons of high vector population is another task. After the eradication of tsetse flies, mechanically transmitted trypanosomosis (T. vivax and T. evansi) will remain as major problems unless control program is devised along with the current tsetse control activities. To this effect control of T. vivax and T. evansi need to be included in the Pan African trypanosomiasis and tsetse control initiatives. References Abebe, G., Jobre, Y., 1996. Trypanosomiasis: a threat to cattle production in Ethiopia. Rev. Med. Vet. 147 (12), 897–902. Acapovi, G.L., Yao, Y., Goran, E.N., Dia, M.L., Desquesnes, M., 2001. Relative abundance of Tabanids in the Savanna Regions of Coˆte d’Ivoire. Revue d’E´levage et de Me´decine Ve´te´rinaire des pays Tropicaux 54 (2), 109–114. Adeyefa, C.A.O., Dipeolu, O.O., 1986. Ectoparasites of horses in Southwestern Nigeria. Insect Sci. Appl. 7, 511–513. Aklilu, N., 2002. Study on Bovine Trypanosomosis in Selected Sites of Central and Western Tigray, Ethiopia. DVM Thesis, FVM, AAU, Debre Zeit, Ethiopia. Amoudi, M.A., LeClercq, M., 1993. First records and addition of two species Atylotus agricola (Wiedemann) and Hematopota minus-

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